Project Summary Recent years have seen a re-emergence of pertussis as a major public health concern, despite successful mass vaccination programs. Critical pertussis disease in infants is typified by intense bouts of paroxysmal coughing, seizures, and over 150,000 deaths annually. Current antibiotic treatments are only effective if given before the onset of the diseases characteristic cough. For these reasons, there is an urgent need for the development of new treatments. The long-term goal of our group is the development of anti-pertussis therapeutics. Our central hypothesis is that targeting tissue damage and inflammation induced by bacterial virulence factors will have a greater benefit than traditional bactericidal therapies. We have previously demonstrated the potential of sphingosine-1-phosphate receptor (S1PR) agoinsts to reduce pulmonary inflammation and prolong survival. S1PRs regulate a multitude of biological processes throughout the body. The objective of this work is to isolate and understand the beneficial elements of the S1PR agonist-mediated response to inform the diesign of future anti- microbials. We identified peptidoglycan recognition protein 4 (PGLYRP4) an antimicrobial protein, as among the most upregulated genes following S1PR agonism. B. pertussis mediates disease through production of virulence factors. Pertussis toxin (PT) is responsible for the propogation and exacerbation of inflammatory responses. Tracheal cytotoxin (TCT) a peptide structurally equivalent to a monomeric subunit of PGN, causes tissue damage and destruction. We hypothesize that PGLYRP4 targets PT and TCT preventing their activity within the host. PGLYRP4 is required for S1PR agonist mediated disease attenuation. Further, we show that the absence of PGLYRP4 and TCT mediate inflammation and PT expression. In this proposal will characterize PGLYRP4 in B. pertussis inflammation along with the potential for rPGLYRP4 to improve the course of murine disease. We will test the hypothesis that PGLYRP4 elicits its protection against pathology through its actions on PT and TCT. PT expression is controlled by 2CSR elements, and reduced upon S1PR agonist treatment. PGLYRP4 induces 2CSR in other bacteria, we will test the ability of rPGLYRP4 to induce 2CSR in B. pertussis and its impact on PT expression. Further, we will test the hypothesis that PGLYRP4 acts as a TCT sponge or decoy, preventing its activation of host receptors. This work seeks to broaden our understanding of B. pertussis pathogenesis, PGLYRP4 and S1P biology. If our hypotheses are correct we will have identified a novel target for treatment of pertussis and increase our understanding of S1PR mediated antibacterial responses.